Acid sphingomyelinase: identification of nine novel mutations among Italian Niemann Pick type B patients and characterization of in vivo functional in-frame start codon

Niemann Pick disease (NPD) is an autosomal recessive disorder due to the deficit of lysosomal acid sphingomyelinase, which results in intracellular accumulation of sphingomyelin. In the present work we studied 18 patients with NPD type B, including five individuals who presented an intermediate phenotype characterised by different levels of neurological involvement. We identified nine novel mutations in the SMPD1 gene including six single base changes c.2T>G, c.96G>A, c.308T>C, c.674T>C, c.732G>C, c.841G>A (p.M1_W32del, p.W32X, p.L103P, p.L225P, p.W244C, p.A281T) and three frameshift mutations c.100delC, c.565dupC, c.575dupC (p.G34fsX42, p.P189fsX1 and p.P192fsX14). The novel c.2T>G (p.M1_W32del) mutation inactivates the first in-frame translation start site of the SMPD1 gene and in the homozygous status causes NPD type B indicating that in'vivo translation of wild type SMPD1 initiates from the first in-frame ATG. Moreover, the new c.96G>A (p.W32X) introduces a premature stop codon before the second in-frame ATG. As a consequence of either c.2T>G (p.M1_W32del) or c.96G>A (p.W32X), impaired translation from the first in-frame ATG results in a mild NPD-B phenotype instead of the severe phenotype expected for a complete deficiency of the enzyme, suggesting that when the first ATG is not functional, the second initiation codon (ATG33) still produces a fairly functional sphingomyelinase. Analysis of the patients'clinical and molecular data demonstrated that all five patients with the intermediate phenotype carried at least one severe mutation. No association between the onset of pulmonary symptoms and genotype was observed. Finally, the presence of c.96G>A (p.W32X), the most frequent allele among Italian NPD type B population, and c.1799G>C (p.R600P) as compound heterozygotes in association with severe mutations suggested a beneficial effect for both mutations.     

Functional in vitro characterization of 14 SMPD1 mutations identified in Italian patients affected by Niemann Pick Type B disease

Niemann Pick disease (NPD) is an autosomal recessive lysosomal storage disorder caused by the deficient activity of acid sphingomyelinase due to mutations in the SMPD1 gene. We functionally characterized three novel SMPD1 mutations and 11 already reported in the Italian population. Mutant alleles were studied for enzyme activity and protein processing in transiently transfected COS-1 cells. The c.96G>A, c.100delG, c.565dupC, and c.575dupC (p.W32X, p.G34fsX42, p.P189fsX1, and p.P192fs14) alleles expressed no immunoreactive protein and consequently no enzyme activity. In contrast, cells transfected with mutants c.308T>C, c.389T>C, c.674T>C, c.732G>C, c.841G>A, c.1687G>A, c.1799G>A, and c.1799G>C (p.L103P, p.V130A, p.L225P, p.W244C, p.A281T, p.D563Y, p.R600H, p.R600P) expressed protein levels comparable to wild-type ASM expressing cells. Only three of these constructs, c.389T>C, c.1687G>A, and c.1799G>A (p.V130A, p.D563Y, p.R600H), retained residual activity while the other five expressed very low or no enzyme activity. As expected, the c.1669underscore;1670delGT (p.V557fsX18) mutant expressed a completely inactive truncated protein. Interestingly, the c.2T>G (p.M1_W32del) mutant expressed 26.9% of the wild type activity, even though no ASM protein was detected by Western blot analysis, suggesting that the amount of produced enzyme is below detection levels. The results presented in this study are consistent with the wide phenotype variability found in NP type B patients and provide valuable insights into the molecular basis of the disease.     

Identification of 25 new mutations in 40 unrelated Spanish Niemann-Pick type C patients: genotype-phenotype correlations

To better characterize Niemann-Pick type C (NPC) in Spain and improve genetic counselling, molecular analyses were carried out in 40 unrelated Spanish patients. The search identified 70/80 alleles (88%) involving 38 different NPC1 mutations, 26 of which are described for the first time. No patient with NPC2 mutations was identified. The novel NPC1 mutations include 14 amino acid substitutions [R372W (c.1114C>T), P434L (c.1301C>T), C479Y (c.1436G>A), K576R (c.1727G>A), V727F (c.2179G>T), M754K (c.2261T>A), S865L (c.2594C>T), A926T (c.2776G>A), D948H (c.2842G>C), V959E (c.2876T>A), T1036K (c.3107C>A), T1066N (c.3197C>A), N1156I (c.3467A>T) and F1224L (c.3672C>G)], four stop codon [W260X (c.780G>A), S425X (c.1274C>A), C645X (c.1935T>A) and R1059X (c.3175C>T)], two donor splice-site mutations [IVS7+1G>A (g.31432G>A) and IVS21+2insG (g.51871insG)], one in-frame mutation [N961_F966delinsS (c.2882del16bpins1bp)] and five frameshift mutations [V299fsX8 (c.895insT), A558fsX11 (c.1673insG), C778fsX10 (c.2334insT), G993fsX3 (c.2973_78delG) and F1221fsX20 (c.3662delT)]. We also identified three novel changes [V562V (c.1686G>A), A580A (c.1740C>G) and A1187A (c.3561G>T)] in three independent NPC patients and five polymorphisms that have been described previously. The combination of these polymorphisms gave rise to the establishment of different haplotypes. Linkage disequilibrium was detected between mutations C177Y and G993fsX3 and specific haplotypes, suggesting a unique origin for these mutations. In contrast, I1061T mutation showed at least two different origins. The most prevalent mutations in Spanish patients were I1061T, Q775P, C177Y and P1007A (10, 7, 7 and 5% of alleles, respectively). Our data in homozygous patients indicate that the Q775P mutation correlates with a severe infantile neurological form and the C177Y mutation with a late infantile clinical phenotype.     

Molecular analysis of Bruton’s tyrosine kinase gene in Spain

Mutations in Bruton’s tyrosine kinase (BTK) gene result in X linked agammaglobulinemia (XLA). Using Single Strand Conformation Polymorphism (SSCP) followed by direct sequencing 21 mutations were found in 27 patients with an XLA phenotype from 21 unrelated families. We identified 13 novel and 8 known mutations: seven missense (R288W, R544G, P566S, K430E; K374N, L512P, R544S), 5 nonsense (Q196X, Y361X, L249X, Q612X, Q466X), 2 deletions of one nucleotide (A207fsX216, Q612fsX648), 2 deletion‐insertions (V219fsX227, K218fsX228), one insertion of two nucleotides (S572fsX587) and 4 point mutations in donor/acceptor splice sites (g.IVS1+1G>C, g.IVS6+5G>A, g.IVS10+1G>T, g.IVS13‐1GG>CT). Carrier detection was performed in 18 mothers. Only in one case the mutation was found to be de novo. Additionally, BTK mutations were not found in four patients without family history, but with XLA‐compatible phenotype. Hum Mutat 18:84, 2001. © 2001 Wiley‐Liss, Inc.     

婴儿严重肌阵挛癫痫钠离子通道SCNIA基因突变分析

目的 研究婴儿严重肌阵挛癫痫(severe myoclonic epilepsy of infancy,SMEI)患儿钠离子通道a1亚单位基因(sodium channel al subunit gene,SCNIA)突变筛查及遗传特征.方法 收集SMEI患儿23例及其家系成员的临床资料及外周血DNA,采用PCR扩增和DNA直接测序的方法筛查SCNIA基因的26个外显子的突变.结果 23例SMEI患儿中17例有SCNlA基因突变.基因突变率约为73.9%(17/23),其中8例为错义突变(F90S、I91T、A239T、W952G、T1210K,V1335M、V1390M、G1433E),3例为无义突变(R612X、W768X、w1408X),3例为缺失突变(A395fsX400、L556fsX557、V1778fsX1800),1例为插入突变(Y1241fsX1270),1例为剪切部位突变(IVS10+3A>G),1例为同义突变(K1492K),截断突变约占总突变的47.1%(8/17).13个突变位点(F90S、I91T、T1210K、V1335M、G1433E、R612X、W768X,A395faX400、L556fsx557、V1778fsXl800、Y1241fsXl270、IVS10+3A>G、K1492K)经相关检索未见报道.14例突变已证实为新生突变,其余3例突变尚不能确定突变来源.结论 SCNIA基因是SMEI患儿的主要致病基因,约一半为截断突变.SMEI患儿的SCNIA基因突变无热点,且多为新生突变.     

Molecular analysis in Fabry disease in Spain: fifteen novel GLA mutations and identification of a homozygous female

Fabry disease, an X-linked inborn error of glycosphingolipid catabolism, results from mutations in the alpha-galactosidase A gene (GLA). Here we report molecular studies in 22 unrelated Spanish patients with Fabry disease ( 20 males and two females). Fifteen novel mutations were identified. In addition 7 previously described mutations and two previously reported polymorphisms were detected. The 15 novel mutations comprise: eight missense E48K (c.142G>A), W81S (c.242G>C), D170H (c.508G>C), W226C (c.678G>T), Q279R (c.836A>G), C382Y (c.1145G>A), I407K (c.1220T>A), L414S (c.1241T>C); one nonsense W95X (c.284G>A); one insertion Y216fsX15 (c.646_647insT); two small deletions G346fsX1 (c.1037delG), K426fsX23 (c.1277_1278delAA); one gross deletion comprising exons 5, 6, 7; one complex mutation (insertion and deletion) A368fsX24 (c.1102delGinsTTATAC), and one splice-site mutation IVS4+1G>A (c.639+1G>A). One of the females was found homozygous for Q279R mutation and she presented with the classic phenotype since the age of 8 years, this case extending into women the severe phenotype observed in classically affected males. Mutation analysis provided precise identification for 30 heterozygotes among female relatives and detection of a de novo mutation. The molecular studies on Spanish Fabry patients here reported further contribute to the identification of new mutations in this disease, and allow reliable detection of heterozygotes which has consequences for genetic counselling and for treatment.     

Identification of seven novel SMPD1 mutations causing Niemann–Pick disease types A and B

Niemann–Pick disease (NPD) types A and B are autosomal, recessively inherited, lysosomal storage disorders caused by deficient activity of acid sphingomyelinase (E.C. 3.1.4.12) because of mutations in the sphingomyelin phosphodiesterase‐1 (SMPD1) gene. Here, we present the molecular analysis and clinical characteristics of 15 NPD type A and B patients. Sequencing the SMDP1 gene revealed eight previously described mutations and seven novel mutations including four missense [c.682T>C (p.Cys228Arg), c.1159T>C (p.Cys387Arg), c.1474G>A (p.Gly492Ser), and c.1795C>T (p.Leu599Phe)], one frameshift [c.169delG (p.Ala57Leufs*20)] and two splicing (c.316+1G>T and c.1341delG). The most frequent mutations were p.Arg610del (21%) and p.Gly247Ser (12%). Two patients homozygous for p.Arg610del and initially classified as phenotype B showed different clinical manifestations. Patients homozygous for p.Leu599Phe had phenotype B, and those homozygous for c.1341delG or c.316+1G>T presented phenotype A. The present results provide new insight into genotype/phenotype correlations in NPD and emphasize the difficulty of classifying patients into types A and B, supporting the idea of a continuum between these two classic phenotypes.     

Phenylketonuria mutations in Northern China

Mutation spectrum of phenylalanine hydroxylase (PAH) gene in patients with phenylketonuria (PKU) in Northern China is described with a discussion on genotype-phenotype correlation. By using PCR/SSCP and DNA sequencing, all exons of PAH gene in the 185 unrelated patients with PKU from Northern China were studied. A total of 70 different mutations, including 42 missense, 12 splice, 7 nonsense, 5 deletion, 3 insertion, and 1 silence/splice mutations, were detected in 349/370 mutant alleles (94.3%). Deletion, insertion, and frameshift mutations were found for the first time in China PKU patients. The mutations R243Q, EX6-96A>G, R111X, Y356X, and R413P were the prevalent mutations with relative frequencies of 22.2, 11.1, 8.7, 6.5, and 6.5%, respectively. Fifteen novel mutations were identified in this study: I38fsX19, IVS4+3G>C, Y154H, R157K, R157I, T200fsX6, Q267H, Q267E, F302fsX39, G346R, S349A, L367L, R400K, IVS12+4A>G, and IVS12+6T>A. Each of them occurs at very low frequency (0.3-1.1%). The mutation spectrum of PKU in Chinese is similar to other Asian populations but significantly different from European populations. Altogether, 70 different mutations are found in 109 genotypes distributed among 185 PKU patients. As shown by the analysis, the predicted residual activity found in the majority of PKU individuals match their in vivo phenotypes, though evidence is also found for both phenotypic inconsistencies among subjects with similar genotypes and discordance between the in vitro and in vivo effects of some mutant alleles. The study enables us to construct a national database in China serving as a valuable tool for genetic counseling and prognostic evaluation of future cases of PKU.     

Characterization of common SMPD1 mutations causing types A and B Niemann-Pick disease and generation of mutation-specific mouse models

Herein we describe detailed characterization of four common mutations (L302P, H421Y, R496L and DeltaR608) within the acid sphingomyelinase (ASM) gene causing types A and B Niemann-Pick disease (NPD). In vitro and in situ enzyme assays revealed marked deficiencies of ASM activity in NPD cell lines homoallelic for each mutation, although Western blotting and fluorescent microscopy showed that the mutant ASM polypeptides were expressed at normal levels and trafficked to lysosomes. Co-immunoprecipitation of the polypeptides with the ER chaperone, BiP, confirmed these findings, as did in vitro expression of the mutant cDNAs in reticulocyte lysates. We further developed a computer assisted, three-dimensional model of human ASM based on homologies to known proteins, and used this model to map each NPD mutation in relation to putative substrate binding, hydrolysis and zinc-binding domains. Lastly, we generated transgenic mice expressing the R496L and DeltaR608 mutations on the complete ASM knock-out background (ASMKO), and established breeding colonies for the future evaluation of enzyme enhancement therapies. Analysis of these mice demonstrated that the mutant ASM transgenes were expressed at high levels in the brain, and in the case of the DeltaR608 mutation, produced residual ASM activity that was significantly above the ASMKO background.     

Identification of four novel PMM2 mutations in congenital disorders of glycosylation (CDG) Ia French patients

We screened 11 unrelated French patients with congenital disorders of glycosylation (CDG) Ia for PMM2 mutations. Twenty one missense mutations on the 22 chromosomes (95%) including four novel mutations were identified: C9Y (G26A) in exon 1, L32R (TA95GC) in exon 2, and T226S (C677G) and C241S (G722C) in exon 8. We studied the PMM activity of these four novel mutant proteins and of the R141H mutant protein in an E coli expression system. The T226S, C9Y, L32R, and C241S mutant proteins have decreased specific activity (23 to 41% of normal), are all more or less thermolabile, and R141H has no detectable activity. Our results indicate that the new mutations identified here are less severe than the inactive R141H mutant protein, conferring residual PMM activity compatible with life.


Keywords: CDG; phosphomannomutase; PMM2 mutations     

A rare lysosomal enzyme gene SMPD1 variant (p.R591C) associates with Parkinson's disease

To investigate the role of mutations in the sphingomyelin phosphodiesterase (SMPD1) gene in Parkinson's disease (PD) we sequenced all the exons of this gene in 198 Chinese PD cases and matched healthy control subjects. We identified 4 rare variants in SMPD1 (p.P332R, p.Y500H, p.P533L, and p.R591C) that were present only in cases and not in control subjects. Interestingly, 2 of these variants were previously reported in Chinese Niemann-Pick disease patients. Next, we genotyped these variants in another 806 PD cases and 7481 control subjects. We identified a novel, rare SMPD1 variant (p.R591C) which increased the risk of PD (p = 0.009).     

Kallmann syndrome: 14 novel mutations in KAL1 and FGFR1 (KAL2)

Kallmann syndrome (KAL) combines hypogonadotropic hypogonadism and anosmia. Hypogonadism is due to Gonadotropin Releasing Hormone (GnRH) deficiency and anosmia is related to hypoplasia of the olfactory bulbs. Occasional symptoms include renal agenesis, bimanual synkinesia, cleft lip palate, dental agenesis. KAL is genetically heterogeneous and two genes have so far been identified, namely KAL1 (Xp22.3) and FGFR1/KAL2 (8p12), which underlie the X chromosome‐linked form and an autosomal dominant form of the disease, respectively. We studied a cohort of 98 unrelated Caucasian KAL patients. We identified KAL1 mutations in 14 patients, of which 7 (c.3G>A (p.M1?), g.IVS1+1G>T, c.570_571insA (p.R191fsX14), c.784G>C (p.R262P), c.958G>T (p.E320X), c.1651_1654delinsAGCT (p.P551_E552delinsSX), c.1711T>A (p.W571R)) have not been previously reported. In addition, we found FGFR1 mutations in 7 patients, namely c.303G>A (p.V102I), C.385A>C (p.D129A), c.810G>A (p.V273M), c.1093_1094delAG (p.R365fsX41), c.1561G>A (p.A520T), c.1836_1837insT (p.Y613fsX42), c.2190C>G (p.Y730X), all of which were novel mutations. In this study, unilateral renal agenesis and bimanual synkinesia were exclusively found associated with KAL1mutations, cleft palate and dental agenesia with FGFR1mutations. © 2004 Wiley‐Liss, Inc.     

Identification of mutations in the GNPTA (MGC4170) gene coding for GlcNAc-phosphotransferase alpha/beta subunits in Korean patients with mucolipidosis type II or type IIIA

Mucolipidosis types II and III are autosomal recessive inherited diseases caused by a deficiency in the lysosomal enzyme N-acetylglucosamine-1 phosphotransferase (GlcNAc-phosphotransferase), which adds phosphate to function as a recognition marker for the uptake and transport of lysosomal enzymes. We investigated mutations in the GNPTA (MGC4170) gene, which codes for the alpha/beta subunits of phosphotransferase, and in the GNPTAG gene, which codes for its gamma subunits in five Korean patients with mucolipidosis type II or IIIA. We identified seven mutations in the GNPTA gene, but none in GNPTAG. The mutations in type II patients included p.Q104X (c.310C>T), p.R1189X (c.3565C>T), p.S1058X (c.3173C>G), p.W894X (c.2681G>A), and p.H1158fsX15 (c.3474_3475delTA), all of which are nonsense or frameshift mutations. However, a splicing site mutation, IVS13+1G>A (c.2715+1G>A) was detected along with a nonsense or a frameshift mutation (p.R1189X or p.E858fsX3 (c.2574_2575delGA)) in two mucolipidosis type IIIA patients. This report shows that mutations in the GNPTA gene coding for the alpha/beta subunits of phosphotransferase, and not mutations in the GNPTAG gene, account for most of the genetic mutations found in Korean patients with mucolipidosis type II or IIIA.     

Mutations of carnitine palmitoyltransferase II (CPT II) in Japanese patients with CPT II deficiency

Carnitine palmitoyltransferase II (CPT II) deficiency is an inherited disorder involving β-oxidation of long-chain fatty acids. CPT II deficiency is a wide-spectrum disorder that includes a lethal neonatal form, an infantile form, and an adult-onset form. However, the ethnic characteristics and the relationship between genotype and clinical manifestation are not well understood. We investigated three non-consanguineous Japanese patients with CPT II deficiency and examined cell lines from 4 unrelated patients and 50 healthy donors. The CPT 2 gene was typed by direct DNA sequencing of polymerase chain reaction-amplified gene products. Case 1 (infantile form) was heterozygous for a phenylalanine to tyrosine substitution at position 383 (p.F383Y) and a novel valine to leucine substitution at 605 (p.V605L). Cases 2, 4, and 5 (infantile form) and case 3 (adult-onset form) were heterozygous for a single mutation at F383Y. Case 6 (adult-onset form) was compound heterozygous at the CPT 2 locus, with deletion of cytosine and thymine at residue 408, resulting in a stop signal at 420 (p.Y408fsX420), and an arginine to cysteine substitution at position 631 (p.R631C). Case 7 (adult-onset form) was homozygous for the p.F383Y mutation. In conclusion, we identified p.F383Y mutations in six of seven patients with CPT II deficiency and two novel variants of the coding gene: p.Y408fsX420 and p.V605L. These mutations differ from those in Caucasian patients, who commonly harbor p.S113L, p.P50H, and p.Q413fsX449 mutations; therefore, our data and those of other Japanese groups suggest that the p.F383Y mutation is significant in Japanese patients with CPT II deficiency.     

Type I Gaucher's disease with homozygous R463C mutation without neurological involvement

Gaucher's disease is an inherited glycolipid storage disorder, caused by a deficiency of the catabolic enzyme glucocerebrosidase. Frequently, N370S and L444P of mutations are observed. R463C (i.e., 1504C-->T) mutation may predict neurological involvement in Gaucher's disease. We report a 36-year-old Turkish man with type I Gaucher's disease with homozygous R463C mutation without neurological involvement.     

Gaucher disease in Romanian patients: incidence of the most common mutations and phenotypic manifestations

Gaucher disease (GD) is an inherited glycolipid storage disorder resulting from the deficiency of glucocerebrosidase. It is the most frequent lysosomal storage disease in Romania, accounting for 70% of all lysosomal disorders diagnosed since 1997 in this country. The prevalence of six common mutations (N370S, L444P, R463C, 84GG, recNciI and recTL) and their phenotypic impact were studied in 20 type 1 GD patients of non-Jewish origin. Mutation analysis identified 77.8% of the GD alleles. The N370S mutation had the highest prevalence (50%), followed by the L444P (22.2%) and the recNciI (5.6%) alleles. Mutations R463C, 84GG and recTL have not been found in our patients. Rare or novel mutations likely accounted for 22.2% of the disease-producing uncharacterised alleles. Our study indicates a high prevalence of type 1 among Romanian GD patients. Clinical phenotype and disease severity were evaluated according to the standardised severity score index. Genotype-phenotype correlations were similar to those reported for other Caucasian non-Jewish populations. The absence of neuronopathic disease in patients presenting at least one copy of the N370S allele was confirmed, but the relative mildness of N370S homozygotes was not a constant feature among our patients. The presence of the L444P or of uncharacterised sporadic mutations was always associated with severe clinical manifestations, even in compound heterozygotes with the N370S allele. A large degree of phenotypic variability was observed in patients displaying the same genotype. The particularities of genotype-phenotype correlations may suggest the impact of other genetic or non-genetic factors on the clinical picture.     

Fabry disease: 45 novel mutations in the alpha-galactosidase A gene causing the classical phenotype

The nature of the molecular lesions in the alpha-galactosidase A (alpha-Gal A) gene causing Fabry disease was determined in 50 unrelated families with the classic phenotype of this X-linked recessive lysosomal storage disease. Genomic DNA was isolated from affected males or obligate carrier females, and the entire alpha-Gal A coding region as well as the flanking and intronic sequences were analyzed by PCR amplification and automated sequencing. Forty-five new mutations were identified including 38 single base substitutions (32 missense and four nonsense) and nine gene rearrangements: MIR, M42T, G43D, G43V, H46Y, F50C, L68F, G132R, T141I, Y152X, K168R, G183S, V199M, P205R, Y207S, Q221X, C223R, C223Y, D234Y, G271C, A288P, P293A, R301G, I303N, I317T, E341D, P362L, R363C, R363H, G373D, I384N, T385P, Q396X, E398K, S401X, P409A, g7325insC, g7384del13, g8341delG, g8391del4/ins3, g10511delTAGT, g10704delACAG, g11019insG, g11021insG, and g11048delAGG. In the remaining five Fabry families, four previously reported mutations were detected (W81X, R112C, g11011delTC, and g11050delGAG) of which the R112C substitution was found in two families who were unrelated by haplotyping. These studies further define the heterogeneity of mutations in the alpha-Gal A gene causing the classical Fabry disease phenotype, and permit precise carrier detection and prenatal diagnosis in these families.     

Molecular basis of familial hypercholesterolemia in Brazil: Identification of seven novel LDLR gene mutations

Low-density lipoprotein receptor (LDLR) gene mutations cause familial hypercholesterol-emia (FH), one of the most common single gene disorders. The spectrum of LDLR mutations in Brazil is not known. The aim of this study was the characterization of LDLR mutations in 35 unrelated Brazilian patients with heterozygous FH. The promoter region, the 18 exons and the flanking intron sequences of the LDLR gene were screened by PCR-SSCP analysis and by DNA sequencing. In addition, we have screened the apolipoprotein B gene (APOB) for known mutations (R3500Q and R3531C) that cause Familial defective apo B-100 (FDB) by PCR-RFLP procedure. We found two nonsense (E92X and C371X) and six missense LDLR mutations (R236W, G322S, G352D, A370T, C675W and C677Y), that were previously described in FH patients from other populations. We also found five novel missense [G(-20)R, T476P, V503G, D580H and S652R] and two novel frame shift LDLR mutations (FsR757 and FsS828). Four patients were found to carry two different mutations in the LDLR gene: G352D and A370T (one patient), S652R and C675W (one patient) and T476P and V503G (two patients). APOB mutations were not found. These findings demonstrate that there is a broad spectrum of mutations in the LDLR gene in FH individuals from Brazil.